The prion protein (PrP) is a brain glycoprotein which has been implicated in the pathogenesis of scrapie, a progressive degenerative neurologic disease of animals which resembles certain diseases of humans (1). Our studies on the biogenesis of PrP in cell-free systems have found a novel mechanism by which the topology of nascent PrP can be regulated at the endoplasmic reticulum to generate either a transmembrane or a secretory form of the nascent polypeptide (2). We have hypothesized that alteration of the topology of PrP may be a central feature of the disease process in scrapie. In support of this, a striking correlation between mutations which affect PrP topology and those which confer disease has been observed (3, 4). Recently, we have discovered that, in Xenopus oocytes, PrP topology is restricted to the secretory form, even though oocytes can recognize information within the sequence of PrP to direct both transmembrane and secretory forms when those coding regions are engineered into the coding regions of other proteins or when the native structure of PrP is disrupted by insertions. Thus, features of PrP distinct from those which direct topology in cell-free systems, appear to restrict the polypeptide to one of its two possible topologic phenotypes in living cells. Here we propose to study this phenomenon of topologic restriction as part of a program to understand cellular responses to PrP. First, we shall characterize this restriction and determine its molecular basis. Second, using topologic restriction as a physiologic parameter, we shall assay the cellular response to both exogenous PrP presentation and endogenous PrP expression. We will determine if topologic restriction is controlled by signalling events from the cell surface and attempt to develop a cell-free system displaying its characteristics. Finally, we shall determine if it can be terminated by new gene products produced during the progression of scrapie. If this proves to be the case, we will clone the responsible genes. In this way, a new cell biologic observation involving PrP will be characterized and assimilated with existing knowledge on both protein trafficking and scrapie pathogenesis. Such investigation in detail of the cellular response to PrP should provide important information relevant to host defense against prion disease. Eventually, this approach may allow molecular insight into possible ways in which biogenesis of the disease-causing phenotype of PrP can be prevented, altered, countered or controlled.